SUMMARY/ABSTRACT The broad impact of this SBIR phase I project is to develop and commercialize a novel biophysical instrument that will greatly enhance researchers? capability to identify better drug candidates for molecular medicine. Significance: Biophysical analysis of molecular interactions is not only essential for disease-related biomedical research, but has also become increasingly popular for drug development, especially in Fragment-based Drug Discovery against challenging targets such as protein-protein interaction interfaces. However, current biophysical techniques (e.g. SPR, ITC and fluorescence-based methods) offer only limited throughput, and mostly require extra pre-treatment steps (labeling or immobilization) that likely result in signal artifacts and enormous effort on assay optimization. At present, significant tradeoff seems inevitable among the throughput, accuracy and sensitivity when choosing the method of biophysical screening, and the result is a prolonged screening process with likely suboptimal lead compound, which increases the chance of failures in downstream drug development. Innovation: We have developed a new instrument that offers unparalleled speed, accuracy and convenience in molecular interaction analysis, without the need for labeling or immobilization. It is based on Transient Induced Molecular Electronic Spectroscopy (TIMES), a novel technology that records transient electrical signal induced by proteins? configuration-specific charge distribution which is highly sensitive to ligand binding. The device also leverages benefits of the microfluidic system and realizes accurate measurement at minimal sample consumption in a high-throughput fashion. It is a universal tool for rapid assessment of biomolecular binding affinities regardless of size, structure or charge status, fitting strong commercial needs from both biophysical labs (for daily protein research) and pharmaceutical industry (for high throughput screening). Preliminary Data: We have demonstrated by using a prototype device that TIMES accurately reproduced affinity values of five classic protein-ligand pairs with diverse molecular properties. Specific Aims: In Specific Aim 1, we will optimize the current device design and algorithm to further improve signal-to-noise ratio and its versatility. Several proposed strategies will be tested and optimal improvements will be incorporated in developing a robust commercial prototype. In Specific Aim 2, we will demonstrate TIMES? applicability for drug discovery by applying it to high-impact protein-ligand interaction pairs, especially those for fragment based drug discovery and conformation-specific modulators, and comparing its accuracy and efficiency with existing technologies in parallel. In Specific Aim 3, we will demonstrate the scalability of TIMES for high-throughput settings by constructing a new design with multiplied microfluidic channels and embedded valve system to achieve a sample throughput that is >10 times higher than current techniques. Together, these studies will firmly establish the commercial utility and feasibility of TIMES instrument for both biophysical research and drug discovery.